Project Summary Precise wiring of the nervous system depends on the guided growth of axonal fibers to their specific targets for synaptic connections. Axon pathfinding depends on the motile growth cone at the tip of developing axons, which senses and responds to a variety of extracellular signals to navigate through a complex and changing environment. The proposed study aims to elucidate a novel signal amplification mechanism that enables growth cones to sense shallow gradients of guidance cues to generate highly polarized actin-based motility. The work will take advantage of a well-defined neuronal culture system for growth cone turning assays, sophisticated high-resolution imaging techniques, direct manipulation of intracellular molecules, and molecular and pharmacological manipulation of cellular components. The central hypothesis is that phosphoinositide-based signal amplification enables the growth cone to sense shallow gradients of guidance cues to generate directional responses. The proposed study will further investigate the molecular mechanisms underlying the phosphoinositide-based signal amplification. The goal is to gain a mechanistic understanding on how a growth cone translates spatiotemporally distributed extracellular cues to distinct directional responses, leading to the formation of specific synaptic connections. The cell?s ability to sense the environment and to determine the direction and proximity of an extracellular stimulus, followed by correct movement, is fundamental for many developmental events including neural development. Directed cell motility also underlies many pathological events, especially cancer-cell metastasis. The proposed study uses nerve growth cones as the model to study the signaling mechanisms that underlie the directional sensing of growth cones during axon pathfinding. The results from this study will not only provide significant insights into the molecular mechanisms of axon guidance, but also extend our knowledge concerning directed cell movement in many physiological and pathological events. Therefore the work is directly relevant to public health.